Interleukin-10 (abbreviated to IL-10 in the following) is a cytokine produced mainly from several types of lymphocytes, such as type 2 helper T-cells (Th2), regulatory T-cells. Though the bioactivity thereof diverges greatly, the characteristic conspicuously varying from other cytokines is that its “inhibitory activity” has become the central issue. IL-10 has inhibitory control of immune function including the production of inflammatory cytokines such as IFN-γ (non-patent reference 1) and exhibits an inhibiting action indirectly towards lymphocytes via monocytic cells. That is, IL-10 exerts an anti-inflammatory effect in human body and maintains immune homeostasis. Its expression in mucous membranes such as the digestive tract is particularly distinguishing (non-patent reference 2). In human genome-wide analysis, the IL-10 signal shows an interrelation with intestinal disorders (non-patent references 3 and 4); the relationship inflammatory in gastrointestinal tract and the anti-inflammatory due to IL-10 is understood to be close and physiological (non-patent reference 5).
In this way, IL-10 is expected to inhibit inflammation and be a treatment drug for autoimmune disease (non-patent reference 6). If IL-10 can be manufactured in large quantities at a low cost using recombinant plant matter, we can expect the development of new use such as administering orally or spraying to mucosa of the nose.
In prior art, the synthesis of recombinant IL-10 is carried out by production using recombinant technology utilizing micro-organisms such as E. coli or mammalian cultured cells (the “method of prior art” as mentioned in the following); but, when mammalian cells are used as parasitic host cells, an enormous expense is involved in order to obtain a large quantity of recombinant IL-10, and the possibility of mixing pathogens such as viruses that infect humans, prion cannot be avoided. Further, the mixing toxicant such as endotoxins becomes a problem with recombinant IL-10 (non-patent reference 7). Particularly in the recombinant IL-10 purified by the method of prior art, the one which is endotoxin-free is not known up to now, and although the above-mentioned anti-inflammatory effect of IL-10 is known, this has not actually been applied in treatments of anti-inflammatory effect.
To solve these problems, the present inventors have considered a method for the expression of IL-10 in rice plants and the purification thereof. Examples in which humans are infected by plant viruses are not known, particularly rice plant is used in food for elderly by human, so as its safety has been confirmed. Accordingly, the risk of the mixing toxicant such as endotoxins is expected to be extremely low. Further, after recombinant rice plants expressing IL-10 are created once, the phylesis can be maintained by only cultivating these and recombinant IL-10 can be produced at extremely low cost in need of the cost for the cultivation only (patent reference 1 and non-patent references 8-9).